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Yumlu, S.* ; Stumm, J. ; Bashir, S.* ; Dreyer, A.K.* ; Lisowski, P.* ; Danner, E.* ; Kühn, R.*

Gene editing and clonal isolation of human induced pluripotent stem cells using CRISPR/Cas9.

Methods 121-122, 29-44 (2017)
Postprint DOI PMC
Open Access Green
Human induced pluripotent stem cells (hiPSCs) represent an ideal in vitro platform to study human genetics and biology. The recent advent of programmable nucleases makes also the human genome amenable to experimental genetics through either the correction of mutations in patient-derived iPSC lines or the de novo introduction of mutations into otherwise healthy iPSCs. The production of specific and sometimes complex genotypes in multiple cell lines requires efficient and streamlined gene editing technologies. In this article we provide protocols for gene editing in hiPSCs. We presently achieve high rates of gene editing at up to three loci using a modified iCRISPR system. This system includes a doxycycline inducible Cas9 and sgRNA/reporter plasmids for the enrichment of transfected cells by fluorescence-activated cell sorting (FACS). Here we cover the selection of target sites, vector construction, transfection, and isolation and genotyping of modified hiPSC clones.
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Publikationstyp Artikel: Journalartikel
Dokumenttyp Wissenschaftlicher Artikel
Korrespondenzautor
Schlagwörter Crispr ; Cas9 ; Gene Editing ; Knockin ; Knockout ; Pluripotent Stem Cells; Human Ips Cells; Crispr-cas9 Nucleases; Genome; Mutations; Specificity; Expression; Efficiency; Talens; Cas9; Dna
ISSN (print) / ISBN 1046-2023
e-ISSN 1095-9130
Zeitschrift Methods
Quellenangaben Band: 121-122, Heft: , Seiten: 29-44 Artikelnummer: , Supplement: ,
Verlag Elsevier
Verlagsort Amsterdam [u.a.]
Nichtpatentliteratur Publikationen
Begutachtungsstatus Peer reviewed
Institut(e) Institute of Developmental Genetics (IDG)